DESCRIPTION (provided by applicant): Cancer, one of the leading causes of death in the United States, is often associated with defects in the major DNA repair systems critical for maintaining genomic integrity. Dysregulation of DNA repair and accumulation of DNA damage is also implicated in the aging process. Base excision repair (BER) is the main line of defense against the most common and mutagenic forms of spontaneous and induced DNA damage. In eukaryotes, many BER proteins are shared by nuclei and mitochondria. While the biochemical operational steps of this important repair pathway have been described in detail, virtually nothing is known about the mechanisms that regulate BER protein localization to the nucleus and/or mitochondria. Organelle-specific DNA damage signals may direct these low-abundance proteins to the appropriate compartment when needed. Since DNA repair and localization mechanisms are highly conserved, the powerful model system Saccharomyces cerevisiae is well-suited for studying both protein localization and DNA repair. An S. cerevisiae BER glycosylase which removes oxidized pyrimidines and is shared by nuclei and mitochondria, Ntg1, appears to be regulated through dynamic localization. I hypothesize that dynamic localization is a general mode of regulation for BER and that DNA repair intermediates produced in the early steps of the BER pathway (e.g. abasic sites) are involved in signaling the presence of DNA damage. This hypothesis will be addressed through two specific aims. Aim 1 will investigate the generality of dynamic compartmentalization by examining the localization of GFP-tagged BER proteins (Ung1, Ogg1, Apn1) in response to preferential genotoxic stress to nuclei or mitochondria. Aim 2 will examine the localization of Ntg1-GFP in cells which have been genetically manipulated to have compartment- specific increases of early BER intermediates to determine whether these intermediates play a role in signaling for dynamic localization. Our long-term goal is to elucidate the signaling pathways that respond to the presence of DNA damage in nuclei or mitochondria, determine how the signals exert their effects, and understand how dysregulation of BER can lead to disease. Furthermore, delineating these signaling pathways will provide valuable insights into the general regulation of nucleomitochondrial signaling and protein localization.

Our biological blueprint, DNA, can be damaged through various means over the natural course of life, but cells make proteins to repair this damage and prevent the development of serious diseases such as cancer and other degenerative disorders. In order to carry out repair, these proteins must be able to get from where they are to the DNA-containing compartments when needed. The proposed research will help us to better understand how DNA repair is regulated - an area which is currently poorly studied yet is critical for comprehending the progression of these diseases - and can potentially provide targets for the development of new anticancer drugs.